摘要
目的:观察DNA去甲基化药物5-杂氮-2′-脱氧胞苷(5-aza-CdR)对前列腺癌细胞株PC3抑癌基因GSTP1和RASSF1A转录改变以及细胞增殖活性的影响。方法:用不同浓度5-aza-CdR(2、5、10μmol/L)处理前列腺癌细胞株PC3,利用甲基化特异性PCR(MSP)法检测用药前PC3细胞株GSTP1、RASSF1A启动子甲基化状态;采用实时荧光定量RT-PCR法检测用药过程中GSTP1和RASSF1A mRNA转录改变;用MTT法和流式细胞术检测用药前后PC3肿瘤细胞增殖活性改变。结果:GSTP1、RASSF1A启动子区域出现甲基化现象。与对照组相比,药物组培养24h后GSTP1和RASSF1A mRNA表达未发生明显改变;培养48h后5、10μmol/L药物组GSTP1和RASSF1A mRNA表达出现上调;72h后各浓度药物组均检测出两基因mRNA表达升高(P<0.05)。药物组培养24和48h未出现明显的细胞增殖抑制现象和细胞周期改变;培养72h后,药物组细胞增殖抑制显著(P<0.05),细胞周期改变明显(P<0.05),阻滞于G0/G1期。结论:GSTP1和RASSF1A基因在PC3前列腺癌细胞系中的失表达可能与其启动子CpG岛高甲基化有关;5-aza-CdR能在一定程度上抑制PC3细胞增殖,干扰细胞周期,提高GSTP1和RASSF1A的转录水平。
Objective：To observe the effects of 5-aza-2＇-deoxycitydine （5-aza-CdR） on the proliferation and transcription of tumor suppressor gene GSTP1 and RASSF1A in prostate cancer cell line PC3. Methods： The status of 5＇CpG island methylation of RASSF1A and GSTP1 genes in PC3 was analyzed by methylation specific PCR （MSP） before treatment with 5-aza-CdR. RASSF1A and GSTP1 mRNA were quantified by real-time PCR during the demethylation process by 5-aza-CdR. MTT assay and flow cytometry were used to examine the proliferative activity of PC3 cells before and after 5-aza-CdR treatment. Results： The 5＇ CpG island methylation of RASSF1A and GSTP1 genes were detected in human prostate cancer cell line PC3. Compared with control group, RASSF1A and GSTP1 mRNA expression had no significant change 24 h after culture with 5-aza-CdR; their expression was up-regulated 48 h after cultured with 5 aza-CdR, with significant difference found between 5μmol/L and 10μmol/L 5-aza-CdR groups. Compared with control group, the expression of RASSF1A and GSTP1 mRNA was significantly increased 72 h after cultured with all concentrations of 5-aza- CdR. MTT assay and cell cycle examination indicated that exposure to 5-aza-CdR for 24 h and 48 h resulted in no obvious growth inhibition and cell cycle change; exposure to 5 aza-CdR for 72 h induced significant growth inhibition （P〈0.05） and cell cycle change （P〈0.05）; and cells were arrested at G0/G1 phase. Conclusion： The 5＇CpG island methylation of RASSF1A and GSTP1 genes is probably responsible for RASSF1A and GSTP1 silencing in PC3 cells. 5-aza-CdR can inhibit the proliferation of PC3 cells, disturb the cell cycle, and elevate transcription of GSTP1 and RASSF1A.
出处
《第二军医大学学报》
CAS
CSCD
北大核心
2009年第3期256-259,共4页
Academic Journal of Second Military Medical University
基金
上海市卫生局科研课题[200646(85)]~~
